Purpose:
Mutations in the RPGR gene cause one of the most common and severe form of X-linked RP in man. AAV-mediated gene transfer of human RPGR 1-ORF15 cDNA rescues photoreceptors (PR) in a canine model of XLRP (c.1084-1085delGA) when delivered at 5 wks of age near the onset of degeneration and evaluated between 21 and 36 wks of age. We now report on the long-term follow-up of RPGR mutant dogs treated with the same viral construct at early, mid and late disease stages.

Methods:
An AAV2/5 vector construct carrying full-length human RPGR 1-ORF15 cDNA under control of a hIRBP promoter was injected subretinally. Early-stage disease (n=7 eyes; age=5 wks; ONL near normal thickness) was treated with vector titers ranging from 0.05 to 15.1 x 1011 vg/ml (70 μl). Mid-stage (n=3 eyes; age=12 wks; ONL=~60% of normal), and late stage disease (n=3 eyes; age=26 wks: ONL=~50-40% of normal) were treated with 150 μl at 1.51x1011 vg/ml. Contra-lateral eyes (n=7) were either injected with BSS, or a similar dose of viral construct intravitreally. All dogs were followed to ages ranging from 36 to 95 wks of age. PR structure and function was assessed by SD-OCT and ERG.

Results:
In early-stage treated eyes, ONL rescue was confirmed at 1.51-15.1x1011 vg/ml titers; at 0.05-0.15x1011 vg/ml there was partial or no efficacy. Thus, a titer of 1.51x1011 vg/ml was used in mid- and late-stage disease. In mid-stage disease, better preserved ONL thickness was detectable on OCT by 20-30 wks post-treatment in the treated area. The difference between treated and untreated regions became magnified as the untreated ONL progressively thinned. In late-stage treated eyes, there was small but detectable preservation of ONL thickness; quantitative analyses supported positive modification of the natural history of XLPRA2 disease progression by gene therapy. No ONL preservation was seen in BSS control eyes. ERG function was greater in treated than in control eyes for all 3 groups using rod and mixed rod-cone dark-adapted b-waves as well as light adapted 29 Hz flicker cone responses.

Conclusions:
Stable and long-term PR rescue (up to 90 wks post-Tx) was achieved with subretinal RPGR gene augmentation therapy delivered at early, mid and late stages of disease. These preclinical results would support gene therapy intervention in retinal areas of RPGR-XLRP patients that retain ~50% of normal ONL thickness.